Discipline: Biological Sciences
Subcategory: Cell and Molecular Biology
Session: 2
Room: Exhibit Hall
Manie Brema - Truman State University
Co-Author(s): Sarah Colijn, PhD, Washington University in Saint Louis, Saint Louis, MO; Amber Stratman, PhD, Washington University in Saint Louis, Saint Louis, MO
Congenital heart defects (CHDs) are common birth defects that affect 1% of births per year in the United States. CHDs occur when the heart develops abnormally affecting its function and may present as a malformation in the blood vessels. Most causes of CHDs remain unknown, so understanding novel regulators of vascular stability during development may be important in understanding drivers of CHDs. During development, vascular stabilization helps strengthen and support the blood vessel with three main components: the endothelial cells that form the vessel wall, the basement membrane that keeps the endothelial cells in place, and the mural cells (MCs), also known as smooth muscle cells, that provide barrier and strength to the blood vessel. Interestingly, during development MCs are preferentially recruited to the arteries over veins. However, the mechanisms of how this choice is made still need to be defined. From preliminary data, using zebrafish, we found that blood flow plays a role in the recruitment of MCs. We hypothesize that primary cilia on endothelial cells serve as mechanosensors for blood flow and influence how MCs get recruited differently onto arteries and veins. The objective of our study is to characterize the vascular phenotypes in cilia mutant zebrafish embryos. In particular, we aim to assess the gross morphology of zebrafish mutants, visualize the cilia in mutant and wildtype zebrafish, and determine if mutant cilia affect the rate of blood flow. Using microscopy, we observed cilia mutant embryos and found that all mutants had curved bodies compared to the controls with straight bodies. We also noticed severe pericardial edema and intracranial hemorrhage in mutant embryos, which may indicate a vascular stability defect. We observed that as cilia-deficient zebrafish embryos aged, there was an increase in pericardial edema, though the intracranial hemorrhage eventually resolved compared to non-mutant siblings. To confirm that the cilia mutants had fewer cilia, we used confocal microscopy to view the primary cilia on the vessels, counted the number of cilia in control versus mutant animals, and found that the mutants had fewer cilia compared to the control. We also calculated the heartbeat of the zebrafish embryos and found that there was no significant difference between mutants and controls, indicating that cilia do not change the blood flow rate itself. Based on our data, we conclude that primary cilia may play a role in vascular stability. This work is significant because discovering the mechanisms behind vascular stability can help us understand and combat developmental cardiovascular disorders like CHDs. In the future, we will look at the role of primary cilia in basement membrane deposition and MC recruitment by analyzing control vs mutant embryos.
Funder Acknowledgement(s): Cell Biology and Physiology Department at Washington Unverisuth Saint Louis School of Medicine, The Genome Institue at Washington University Saint Louis, Opportunities in Genomic Research Program
Faculty Advisor: Sarah Colijn, PhD, colijn@wustl.edu
Role: For my research project, I had to genotype the zebrafish with the ift172 cilia deficient protein. This allows me to detect what zebrafish had the mutation and breed them to get the eggs. Then I incubated the eggs for around 42 to 72 hours to phenotype the embryos. I also characterize the phenotype of the zebrafish embryos that had cilia mutants. I had to keep track of the amount of curved and straight bodies in each clutch, and from these clutches, I had to look at their phenotype from 42 to 72 hours post-fertilization. I also used the confocal microscope to observe the number of cilia present in the vessel of both mutant and control. For the cilia, I also had to keep notes of observation of cilia features. This information allows me to see a possible trend with the mutant or control if they have more or fewer cilia. When calculating the heart rate, I used a confocal microscope to record, count, and calculate the heartbeat every 10 seconds